Natural killer (NK) cells play an important function in antitumor and anti-microbial defense, in regulation of lymphoid and hematopoietic cells, in production of a variety of cytokines, and represent the dominant effector cells involved in lymphokine-activated killer cell (LAK) phenomenon. Despite the biological and clinical significance of NK cells, very little attention has been given to defining NK cell lineage and understanding the maturational events in NK cell development. Our proposal is aimed at bridging this gap. It is now possible to study NK cells lineage and development as a result of the recent advances in recombinant DNA technology, molecular approaches and availability of a plethora of monoclonal antibodies, cytokines and genetic probes. The specific aims of our investigations are to study: (1) Generation of NK cells from human bone marrow CD34+ progenitors and their subsets, including hematopoietic stem cells (HSC), using long-term bone marrow culture (LTBMC) systems; (2) Stepwise characterization of NK cell differentiation events in LTBMC and subsequent clonal analysis of NK cell progenitors; LTBMC will be analyzed for: morphology and for acquisition of cytoplasmic and cell surface antigens during maturation, using two and three-color flow cytometry; expression of different forms of NK cell tumor recognition structure (NK-TR), as determined by alternate mRNA splicing, using PCR technique; zeta subunit, using cell-permeabilization procedure; frequency of NK progenitors, using limiting dilution assay; an attempt will be made to prepare monoclonal antibodies against NK cell progenitors. (3) Events in development of NK cell cytotoxic and regulatory functions in LTBMC, including development of NK cell cytotoxic mechanism, acquisition of binding/lytic activity, granzyme and cytolysin systems, ability to produce cytokines and regulate hematopoiesis. This step-wise analysis of NK cell maturation, starting with the most primitive stem cell compartment, will allow us to "map", for the first time, NK differentiation pathway(s). This information will help to answer the questions of NK cell lineage and will be instrumental in determining the maturational pathways involved in acquisition of some of the NK cell biological functions. By "mapping" of normal NK cell matura- tion, it may be possible to detect divergent pathways of differentiation or asynchronous structure/function relationships in cancer patients, especially leukemia.